Beta-galactosidase (beta-D-galactoside galactohydrolase, EC 3.2.1.23) is an enzyme capable of hydrolyzing the disaccharide lactose to its monosaccharide constituents, D-glucose and D-galactose. Beta-galactosidases are found in a large variety of organisms, like mammals, plants, fungi, yeasts, and bacteria. In Nature, beta-galactosidases hydrolyze lactose and other D-galactose-containing carbohydrates. In the industry, beta-galactosidases have been used primarily within the food industry. Beta-galactosidase hydrolysis of lactose and lactose-containing dairy products are used throughout in the dairy industry in the preparation of lactose-free or low-lactose products, which may be consumed by humans suffering from lactose intolerance. Hydrolysis of lactose by beta-galactosidases may also be used in applications where the removal of lactose is required, i.e. prevention of crystallisation of lactose in food and removal of D-galactose moieties in glycosylated proteins. Other applications of beta-galactosidases comprise hydrolysis of lactose into D-galactose and D-glucose with the subsequent modification of the monosaccharides to high value products, like the sweetener D-tagatose (Jorgensen et al. 2004).
Application of beta-galactosidases could be used to produce lactose-free and low-lactose dairy products for lactose intolerant humans.
The major applications for lactose hydrolysis are listed below.                a) Liquid milk. Lactose hydrolysis in liquid milk improves digestibility for lactose intolerant consumers. In flavoured milks, lactose hydrolysis increases sweetness and enhances flavours.        b) Milk powders. Lactose hydrolysed milk powders for dietetic uses, especially for infants with temporary beta-galactosidase deficiency.        c) Fermented milk products. In some cases, lactose hydrolysis in milk used for the manufacture of cheese and yoghurt can increase the rate of acid development and thus reduce processing time.        d) Concentrated milk products. Lactose hydrolysis in concentrated milk products (e.g. sweetened condensed milk, ice cream) prevents crystallisation of lactose.        e) Whey for animal feed. Lactose hydrolysis in whey enables more whey solids to be fed to pigs and cattle and also prevents crystallisation in whey concentrate.        f) Whey. Lactose hydrolysed whey is concentrated to produce a syrup containing 70-75 percent solids. This syrup provides a source of functional whey protein and sweet carbohydrate and is used as a food ingredient in ice cream, bakery and confectionery products.        
The conventional approach in food processing is to carry out the hydrolysis of lactose at 40° C. during approximately four hours.
However, milk or lactose solution as a raw material is a preferable nutrition source for bacteria. As the result, the putrefaction owing to the saprophyte contamination during the treatment is a serious problem in the food production. Thus, the fact is that the conventional beta-galactosidase is of limited use.
Most beta-galactosidases in practical use are active only at temperatures above 20-30° C., temperatures where food spoiling bacteria thrive at best.
Attempts to use thermophilic beta-galactosidases have been used but the products have suffered from off flavours and reduced organoleptic properties due to the heat treatment, and the processes have demanded high energetic costs.
A number of cold-active beta-galactosidases have been described from Arthrobacter (Coker, et al. 2003; Karasová-Lipovová, et al. 2003; Nakagawa et al. 2003; Nakagawa et al. 2006), from Carnobacterium piscicola (Coombs and Brenchley, 1999) and from Pseudoalteromonas (Cieslinski, et al. 2005; Fernandes, et al. 2002; Hoyoux, et al. 2001; Turkiewicz, et al. 2003). Furthermore, Nakagawa et al. (2006) described a cold-active beta-galactosidase from the yeast Guehomyces pullulans. However, the activity of cold-active beta-galactosidases described so far is low at the low temperatures, which is wanted by the dairy industry. The beta-galactosidase from the yeast Guehomyces pullulans had approximately 17% at 0° C. (Nakagawa et al. 2005), the beta-galactosidases from Carnobacterium piscicola BA showed approximately 24% activity at 10° C. (Coombs and Brenchley, 1999) and the enzymes from Pseudoalteromonas isolates showed 39% activity (Fernandes et al. 2002), 22% activity (Cieslinski et al. 2005), and 12% activity (Hoyoux et al. 2001) at 10° C. So far, the beta-galactosidases with highest activity at low temperatures have been isolated from Antarctic Arthrobacter isolates. Karasová-Lipovová, et al. (2003) showed that a psychrotolerant Arthrobacter sp. C2-2 isolate produced beta-galactosidase, which displayed 19% of its maximal activity at 10° C., Coker et al. (2003) described an enzyme from an Antarctic Arthrobacter isolate with approximately 50% at 0° C., and Nakagawa et al. (2003, 2006) described a beta-galactosidase from A. psychrolactophilus F2, which had its temperature optimum at 10° C.
However, the cold-active beta-galactosidase from the Antarctic Arthrobacter was produced in low amounts in native cells and attempts to produce the enzyme recombinantly in E. coli were unsuccessful since about 90% of the enzyme was located in insoluble inclusion bodies (Coker et al. 2003). The cold-active beta-galactosidase from A. psychrolactophilus F2 could be produced heterologously, but had lower activity than the other Arthrobacter beta-galactosidases (Nakagawa et al. 2006).
Therefore, in order to develop a low-temperature process for hydrolysis of lactose there is a need for a novel cold-active beta-galactosidase and a method for producing such enzyme.